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Acta Agronomica Sinica ›› 2021, Vol. 47 ›› Issue (6): 1090-1099.doi: 10.3724/SP.J.1006.2021.04201


Expression analysis of abiotic stress response gene HcWRKY71 in kenaf and transformation of Arabidopsis

LI Hui1,2(), LI De-Fang2,*(), DENG Yong2, PAN Gen2, CHEN An-Guo2, ZHAO Li-Ning2, TANG Hui-Juan2   

  1. 1College of Life and Environment Science, Hunan University of Arts and Science, Changde 415000, Hunan, China
    2Institute of Bast Fiber Crops,Chinese Academy of Agricultural Sciences, Changsha 410205, Hunan, China
  • Received:2019-12-19 Accepted:2020-10-14 Online:2021-06-12 Published:2020-09-08
  • Contact: LI De-Fang E-mail:guangjunmuzi@126.com;18973612200@163.com
  • Supported by:
    The China Agriculture Research System(CARS-19-E07);The Agricultural Science and Technology Innovation Program at the Chinese Academy of Agricultural Science(ASTIP-IBFC03);The Hunan Education Department Project(18C0737);The Doctoral Research Start-up Project of Hunan University of Arts and Sciences(17BSQD13)


WRKY transcription factor plays an important role in plant responded to abiotic stress. In this study, the unigene sequence (CL3883.Contig4) of the transcriptome in kenaf was used as a reference. Primers were designed for PCR amplification. The full length of HCWRKY71 gene was 957 bp obtained by the sanger sequencing. HCWRKY71 gene had an open reading frame length of 957 bp, and encoded a protein containing 318 amino acids, with a conserved functional domain of WRKY, which belonged to WRKY transcription factor II. Under the salt stress, its relative expression level increased with the increase of NaCl concentration; under the drought stress, with the extension of drought time, the relative expression of the HcWRKY71 gene first decreased, then increased and finally decreased; under the stress of heavy metal cadmium, its expression decreased with the increase of CdCl2 concentration, indicating that the expression of the gene was induced by salt, drought and heavy metal cadmium stress. The gene was transformed into Arabidopsis by Agrobacterium-mediated inflorescence impregnation. It was found that the HcWRKY71 gene improved the salt tolerance of transgenic Arabidopsis seedlings. This laid a solid foundation for further study of stress tolerance mechanism of HcWRKY71 gene.

Key words: kenaf, salt tolerance, drought, heavy metal cadmium, HcWRKY71

Table 1

Primer used in this study"

Primer name
Primer sequences (5°-3°)
Primer usage
Ligation of target gene with expression vector
HcWRKY71-PR TCGGATATGGGCTGTTCTT PCR identification and real-time fluorescence quantitative analysis of transgenic Arabidopsis

Fig. 1

Agarose gel electrophoresis of cDNA full length of HcWRKY71 M: DNA marker 2K Plus II; 1: PCR product."

Fig. 2

Nucleotide sequence and encoded amino acids sequence of HcWRKY71 gene The sequence of WRKYGQK motif is highlighted in red box, that of C2H2 motif (CX4CX23HXH) in black box."

Fig. S1

Conserved domain prediction of HcWRKY71 protein in kenaf"

Fig. S2

Prediction of phosphorylation sites of transcription factor HcWRKY71"

Fig. 3

Amino acid sequence alignment of HcWRKY71 and WRKY71 proteins from other plants The different color indicates that the different levels of the conservation amino acid residues. The blue means amino acids are complete conservation, the pink, cyan, yellow means that the conservation of amino acids is no less than 75%, no less than 50%, no less than 33% and white means no more than 33%."

Fig. 4

Phylogenetic tree of HcWRKY71 and WRKY71 from other plants"

Fig. 5

Expression of HcWRKY71 under salt, drought and cadmium stress A: salt stress; B: drought stress; C: cadmium stress. *, ** mean significant difference at the 0.05 and 0.01 probability levels, respectively. Error bars represent the standard error of each treating group (n = 3)."

Fig. 6

HcWRKY71 expression in different organs under salt, drought and cadmium stress A: salt stress; B: drought stress; C: cadmium stress. *, ** mean significant difference at the 0.05 and 0.01 probability levels, respectively. Error bars represent the standard error of each treating group (n = 3)."

Fig. 7

Identification of transgenic Arabidopsis A: identification of transgenic Arabidopsis by hygromycin; B: PCR identification of transgenic Arabidopsis plant; C: real time quantitative PCR identification. *, ** mean significant difference at the 0.05 and 0.01 probability levels, respectively. Error bars represent the standard error of each treating group (n = 3)."

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